Expression Of Musashi-1 Research Articles

Phenotypic features of rib cartilage cells in funnel-shaped chest deformity

Funnel-shaped deformity of the chest is a serious disease with an unknown etiology and pathogenesis. Pathological changes occurring in the cartilaginous tissue of the ribs are one of the leading mechanisms for the development of chest curvature. Analysis of the phenotypic features of cells isolated from the site of localization of the pathological process will create a more complete picture of the deformation development.Aim of the study was to identify the phenotype of rib cartilage cells in funnel-shaped deformation of the thorax.Material and methods. Rib cartilage cells of children with funnel-shaped chest deformity were cultured up to the 2–3rd passage, then were identified by histochemical and immunohistochemical methods.Results and discussion. The study revealed cells of two different phenotypes. The first type is oval and polygonal chondroblasts expressing a marker of chondrogenic differentiation. The second type is the cells of the neural phenotype with the expression of early neural markers Musashi1 and PAX6 and the neurospecific protein III β-tubulin, as well as the substance Nissl.Conclusions. The analysis of the phenotypic features of rib cartilage cells with funnel-shaped curvature of the chest allowed us to expand the idea of a possible etiological factor in the deformation development.

Musashi-1 Is a Novel Immunohistochemical Marker of Neuroendocrine Carcinoma of the Lung

Small cell lung cancer (SCLC) and large cell neuroendocrine carcinoma (LCNEC) have recently been grouped as lung neuroendocrine carcinomas (NECs). Because these lung NECs are clinically malignant and their treatment strategies differ from those of non-SCLC, the quality of diagnosis has a significant prognostic impact. The diagnosis of LCNEC requires positive immunohistochemical staining with chromogranin A, synaptophysin, and CD56, along with a morphological diagnosis, and insulinoma-associated protein 1 (INSM1) has been proposed as an additional marker but is still not an ideal or better marker. We investigated Musashi-1 as a novel immunohistochemical marker in 42 patients with SCLCs and 44 with LCNECs who underwent lung resection between 1998 and 2020 at our institution. We found Musashi-1 expression in 98% (41/42) SCLC and in 90% (40/44) LCNEC. These findings were similar to CD56 expression and superior to synaptophysin, chromogranin A, and INSM1. Musashi-1 also tended to show more diffuse and intense staining, especially in LCNEC, with more cases staining > 10% than any other existing markers (Musashi-1, 77%; INSM1, 45%; chromogranin A, 34%; synaptophysin, 41%; and CD56, 66%). In conclusion, we identified Musashi-1 as a novel immunohistochemical staining marker to aid in the diagnosis of lung NEC.

FRI291 Musashi1 And Musashi2 Mark Distinct Pituitary Stem Cell Populations

Abstract Disclosure: Y. Rahmatallah: None. J. Banik: None. K. Bronson: None. M.C. MacNicol: None. The cell plastic state of the pituitary is required to meet the changing demands upon pituitary function and may also result in these cells being more vulnerable to malignant transformation. The mechanisms that control pituitary cell plasticity are unknown. The RNA binding protein Musashi (MSI) is expressed in embryonic stem cells, and in cancer stem cells, where MSI functions to promote stem cell plasticity and to oppose premature cell maturation. MSI is also expressed at high levels in adult gonad and adult pituitary. The function of MSI in adult tissues is not fully understood, and the mechanisms that control the pro-proliferative and pro-malignant aspects of MSI function in adult tissues are not known. The two evolutionarily conserved MSI proteins, Musashi1 (MSI1) and Musashi2 (MSI2), are predicted to function redundantly, however, regulatory distinctions have been reported for MSI1 and MSI2. Towards a characterization of MSI1 and MSI2 in pituitary, we analyzed transcriptomics data from public datasets of single-nucleus and single-cell and bulk-tissue gene expression data from physiological and pathophysiological human pituitary tissue. We performed bioinformatics analysis utilizing the single-cell genomics analysis tool, Seurat in R and the pseudotime trajectory analysis tool Monocle and additional genomics and statistics analyses tools in R. We identify the MSI1-expressing pituitary cell population as immature stem cells and the distinct MSI2-expressing pituitary cell population as more mature, committed progenitor cells. Pituitary from human fetal tissue contains multiple distinct MSI1-expressing cell populations while, in contrast, pituitary from human adult tissue contains multiple distinct MSI2-expressing cell populations. Pituitary from human pediatric tissue demonstrates an intermediate state with both distinct MSI1-expressing and distinct MSI2-expressing cell populations. We identify increased MSI1 expression but not increased MSI2 expression in adult pituitary adenoma, and a high level of variation in the MSI1 to MSI2 expression ratio between distinct pituitary adenomas. Based upon this re-analysis of reposited datasets from other pituitary investigators, we hypothesize that the increase that we observe, in MSI1 to MSI2 expression ratio in adult pituitary tumors is consistent with a reversion of mature cells to an earlier immature state. Adult pituitary may contain the mechanisms that regulate MSI2 function but may not be equipped to properly control high levels of re-expressed MSI1. An understanding of the physiological functions and mechanisms of regulation of the distinct MSI proteins in pituitary may provide insight into potential mechanisms that could be recruited to increase physiological pituitary cell growth and differentiation in regenerative therapy and to oppose pathological cell growth in pituitary tumor. Presentation: Friday, June 16, 2023

Musashi-1 regulates cell cycle and confers resistance to cisplatin treatment in Group 3/4 medulloblastomas cells.

Groups (Grp) 3 and 4 are aggressive molecular subgroups of medulloblastoma (MB), with high rates of leptomeningeal dissemination. To date, there is still a paucity of biomarkers for these subtypes of MBs. In this study, we investigated the clinical significance and biological functions of Musashi-1 (MSI1) in Grp3 and Grp4-MBs. First, we assessed the expression profile of MSI1 in 59 primary MB samples (15-WNT, 18-SHH, 9-Grp3, and 17-Grp4 subgroups) by qRT-PCR. MSI1 mRNA expression levels were also validated in an additional public dataset of MBs (GSE85217). The ROC curve was used to validate the diagnostic standards of MSI1 expression. Next, the potential correlated cell-cycle genes were measured by RNA-Seq. Cell cycle, cell viability, and apoptosis were evaluated in a Grp3/Grp4 MB cell line after knockdown of MSI1 and cisplatin treatment. We identified an overexpression of MSI1 with a high accuracy to discriminate Grp3/Grp4-MBs from non-Grp3/Grp4-MBs. We identified that MSI1 knockdown not only triggered transcriptional changes in the cell-cycle pathway, but also affected G2/M phase in vitro, supporting the role of knockdown of MSI1 in cell-cycle arrest. Finally, MSI1 knockdown decreased cell viability and sensitized D283-Med cells to cisplatin treatment by enhancing cell apoptosis. Based on these findings, we suggest that MSI1 modulates cell-cycle progression and may play a role as biomarker for Grp3/Grp4-MBs. In addition, MSI1 knockdown combined with cisplatin may offer a potential strategy to be further explored in Grp3/Grp4-MBs.

Targeted Musashi1 knockdown leads to reduce tau pathology and improves cognitive function in aged P301L mouse model

AbstractBackgroundThe buildup of toxic tau species in the brain is thought to lead to the development of Alzheimer’s disease. While our current understanding of the mechanisms underlying this aggregation is limited, growing evidence has pointed to RNA‐binding proteins (RBPs) playing a role. The dysregulation of RBPs is associated with various neurodegenerative diseases. For example, during amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTLD), dysregulated TDP‐43 accumulates in the brain. Similarly, our recent studies showed that the RBP, Musashi‐1 (MSI1), may co‐aggregate with toxic tau oligomers in Alzheimer’s disease. Clarifying the connection between these two proteins may provide novel insights into the mechanisms behind toxic tau buildup in Alzheimer’s, allowing for new therapeutic targets in treatment development.MethodIn this study, we investigated the role of MSI1 on the aggregation and deposition of tau in aged (18‐months old) P301L mouse models, which express the P301L mutant form of tau. MSI1 expression was knocked down (KD) by performing intracerebroventricular injections of MSI1 antisense oligonucleotides into the hippocampus. The effects of MSI1 KD on tau accumulation and pathology were studied in the hippocampus, cortex, and cerebellum.ResultWestern blot and immunofluorescence (IF) analysis revealed a significant reduction in MSI1 and tau proteins levels and aggregation in the P301L mouse brain, mainly in the hippocampus. Furthermore, brain Co‐IF microscopy and Dot Blot analysis of the PBS‐ and Sarkosyl‐soluble fractions showed that there was a significant reduction of MSI1 and tau oligomeric species in the hippocampus, cortex, and cerebellum in MSI1 KD mice. In addition, phosphorylated tau forms (pSer422, pSer202/Thr205, pThr212/Ser214 and pThr231/Ser235) were significantly decreased in the cortex, cerebellum, and most prominently, in the hippocampus. Increase in synaptic marker Synaptophysin suggesting an increase of synaptogenesis. Behavioral tests, Novel Object Recognition (NOR) and Y‐Maze spontaneous alternations tests, MSI1‐KD mice showed significant improvements.ConclusionThese results suggest that MSI1 plays a major role in tau toxicity, revealing it as a possible target for treating Alzheimer’s disease.

MSI1 Stabilizes MACF1 to Inhibit Apoptosis of MC3T3-E1 Cells Induced by High Glucose and Promote Osteogenic Differentiation Through Wnt/β-Catenin Signaling Pathway.

Diabetes mellitus (DM) affects bone metabolism and causes osteoporosis. Musashi 1 (MSI1), a member of the Musashi family, regulates protein expression by targeting protein mRNA and has been reported to play an important role in osteogenic differentiation. Therefore, this paper attempts to explore the role of MSI1 in diabetic osteoporosis and discussing its specific mechanism. The glucose concentration for high glucose (HG) and control MC3T3-E1 cells were 30 and 5.5mM. MC3T3-E1 cells induced by high glucose (HG) were used to simulate diabetic osteoporosis in vivo. The interaction between MSI1 and microtubule actin crosslinking factor 1 (MACF1) was confirmed by RNA Immunoprecipitation (RIP). The mRNA and protein expressions of MSI1 and MACF1 in MC3T3-E1 cells and HG-induced MC3T3-E1 cells after indicated transfection were tested by Real-time quantitative polymerase chain reaction (RT-qPCR) assay and western blot. After transfection, the proliferation, apoptosis, and osteogenic differentiation of HG-induced MC3T3-E1 cells were detected by cell counting kit (CCK)-8, terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL), alkaline phosphatase (ALP) activity assay, and alizarin red staining. The expression of Wnt/β-catenin signaling pathway-related proteins in HG-induced MC3T3-E1 cells after transfection was detected by western blot. This work shows that MSI1 can combine with MACF1. The expression of MSI1 and MACF1 was increased in HG-induced MC3T3-E1 cells. Upregulation of MSI1 promoted the proliferative and differentiative capabilities, but inhibited the apoptosis of HG-insulted MC3T3-E1 cells, which could be reversed by MACF1 knockdown. MSI1 stabilizes MACF1 to suppress apoptosis and promote osteogenic differentiation in HG-induced MC3T3-E1 cells by inhibiting Wnt/β-catenin signaling pathway.

Bone Marrow Mesenchymal Stem Cell (BMSC) Ameliorates the Damage of Inflammatory Bowel Disease (IBD) Rats Through Regulation of Stromal Cell-Derived Factor 1/C-X-C Chemokine Receptor Type 4 (SDF-1/CXCR4) Axis

This study assesses the effect and mechanism of BMSC in IBD rat. Fifty SDF-grade rats were assigned into divided into NC group, model group, BMSC group, blocking agent group and positive NC group randomly with 10 rates in each group. The histopathological changes of colon tissue, expression of Musashi-1, DAPI, SDF-1 and CXCR4 was measured. There was notable inflammatory cell infiltration in model group and agonist group. The structure of gland was destructed notably with notable-visible phenomenon of hyperemia and edema in colon tissue. They could be improved significantly in positive control group and BMSC group. The necrotic colonic mucosal tissue began to be recovered slowly. The phenomenon of hyperemia and edema was alleviated notably without abnormality in colon tissue in control group. The positive level of Musashi-1 in control group, model group and agonist group was the highest. In conclusion, BMSC could be migrated into colonic damage position and differentiated into intestine stem cells to exert recovery effect on IBD rats. The molecular mechanism might be related with SDF-1/CXCR4 axis.

PMON29 A Comparative Analysis of Musashi-Dependent Control of Mouse and Human Pituitary mRNA Translation

Abstract Specialized cell lineages within the anterior pituitary produce specific hormones which, in turn, regulate diverse biological processes ranging from growth, metabolism, and reproduction to stress. Interestingly, these specialized cell lineages exhibit a high degree of cellular plasticity, readily altering their differentiation state to produce different hormones in response to organismal demand. Underlying molecular mechanisms controlling cellular plasticity in the pituitary are not fully defined but appear to involve post-transcriptional regulation exerted by the Musashi family of sequence-specific RNA binding proteins. Musashi is a bifunctional regulator of target mRNAs and can exert either repression or activation of translation in a context-dependent manner. Our recent work has implicated a role for Musashi-dependent repression of key pituitary mRNAs in the mouse, including a lineage-specific transcription factor (Pou1f1), the gonadotropin releasing hormone receptor (Gnrhr), and two anterior pituitary hormones (Prl and Tshb). In this study, we look to determine the potential relevance of Musashi-dependent mRNA translational control to the human pituitary. Reflecting our findings in the adult mouse, a re-analysis of published human fetal pituitary single cell RNA sequencing data revealed Musashi1 (MSI1) and Musashi2 (MSI2) expression in both stem/progenitor cell populations as well as in all hormone-producing cell lineages of the anterior pituitary. We further report that the human homologs of identified Musashi-target mRNAs in the mouse pituitary also possess Musashi binding elements (MBEs) in their 3' untranslated regions (UTRs). In the case of the human Pou1f1 3' UTR, we were able to verify Musashi-dependent mRNA translational repression in reporter assays as seen with the murine Pou1f1 3' UTR. Interestingly, a germline Pou1f1 mutation identified in human patients, disrupts a consensus MBE in the human Pou1f1 3' UTR and abolishes Musashi-dependent repression. We developed a bioinformatic pipeline to characterize pathogenic human mutations catalogued in the Geno2MP database for disruption or creation of consensus MBEs in pituitary-specific mRNA 3' UTRs. Our findings suggest an evolutionarily conserved role for Musashi in the post-transcriptional regulation of pituitary mRNAs in both mice and humans and identify several human mutations which may perturb Musashi regulation of human pituitary function. Presentation: Monday, June 13, 2022 12:30 p.m. - 2:30 p.m.

Musashi-1 and miR-147 Precursor Interaction Mediates Synergistic Oncogenicity Induced by Co-Infection of Two Avian Retroviruses.

Synergism between avian leukosis virus subgroup J (ALV-J) and reticuloendotheliosis virus (REV) has been reported frequently in co-infected chicken flocks. Although significant progress has been made in understanding the tumorigenesis mechanisms of ALV and REV, how these two simple oncogenic retroviruses induce synergistic oncogenicity remains unclear. In this study, we found that ALV-J and REV synergistically promoted mutual replication, suppressed cellular senescence, and activated epithelial-mesenchymal transition (EMT) in vitro. Mechanistically, structural proteins from ALV-J and REV synergistically activated the expression of Musashi-1(MSI1), which directly targeted pri-miR-147 through its RNA binding site. This inhibited the maturation of miR-147, which relieved the inhibition of NF-κB/KIAA1199/EGFR signaling, thereby suppressing cellular senescence and activating EMT. We revealed a synergistic oncogenicity mechanism induced by ALV-J and REV in vitro. The elucidation of the synergistic oncogenicity of these two simple retroviruses could help in understanding the mechanism of tumorigenesis in ALV-J and REV co-infection and help identify promising molecular targets and key obstacles for the joint control of ALV-J and REV and the development of clinical technologies.

Knockdown of the stem cell marker Musashi-1 inhibits endometrial cancer growth and sensitizes cells to radiation

BackgroundEndometrial carcinoma is the most common gynecological cancer in Europe. Musashi-1 is known to be a key regulator of endometrial cancer stem cells and a negative prognostic marker. In the present study, we aimed to understand growth and gene expression patterns in endometrial carcinoma after Musashi-1 knockdown in vitro and in vivo. Changes in therapeutic resistance were also assessed.MethodsFirst, we performed analyses to understand Musashi-1 expression patterns using The Cancer Genome Atlas database. We then proceeded to assess effects of small interfering RNA-based Musashi-1 targeting in two endometrial carcinoma cell lines, Ishikawa and KLE. After quantifying baseline changes in cell metabolism, we used MTT tests to assess chemotherapy effects and colony formation assays to understand changes in radioresistance. For mechanistic study, we used quantitative polymerase chain reaction (qPCR) and western blotting of key Musashi-1 target genes and compared results to primary tissue database studies. Finally, xenograft experiments in a mouse model helped understand in vivo effects of Musashi-1 knockdown.ResultsMusashi-1 is aberrantly expressed in primary tumor tissues. In vitro, silencing of Musashi-1 resulted in a strong decline in cell proliferation and radioresistance, while chemoresistance remained unchanged. Loss of Musashi-1 led to downregulation of telomerase, DNA-dependent protein kinase, the Notch pathway and overexpression of cyclin-dependent kinase inhibitor p21, the latter of which we identified as a key mediator of Msi-1 knockdown-related anti-proliferative signaling. In vivo, the anti-proliferative effect was confirmed, with Msi-1 knockdown tumors being about 40% reduced in size.ConclusionsMusashi-1 knockdown resulted in a strong decrease in endometrial cancer proliferation and a loss of radioresistance, suggesting therapeutic potential.

Inhibition of Musashi-1 enhances chemotherapeutic sensitivity in gastric cancer patient-derived xenografts.

Musashi-1 (MSI1), a neural RNA-binding protein, is considered a gastric and intestinal stem cell marker. Although the function of MSI1 in gastric cancer has attracted increasing interest, it is not known whether MSI1 can be used as a biomarker to monitor gastric cancer development and response to treatment. Here, the role of MSI1 in the chemotherapeutic sensitivity of gastric cancer was investigated. Patients with high MSI1 levels had poor outcomes, implicating the gene in the development and progression of the disease. We overexpressed and silenced MSI1 in the human gastric cancer cell lines MKN45 and HGC27, finding that knockdown reduced proliferation, invasion, and migration, while promoting apoptosis. A patient-derived xenograft gastric cancer model was constructed in which mice received chemical drugs, si-MSI1, or a drug-si-MSI1 combination. It was found that blocking MSI1 expression reduced gastric cancer drug tolerance. The combination treatment with si-MSI1 was superior to 5F-dUMP and cisplatin, either separately or in combination, indicating that including si-MSI1 was better than drug therapy alone. Transcriptome sequencing analysis showed that MSI1 altered cell cycle regulation and growth signal transduction, including that of blood vessel epicardial substance (BVES). These results suggest that MSI1 reduces the tolerance of gastric cancer to chemical drugs through modulation of MSI1/BVES signaling.

TNF-α up-regulates Nanog by activating NF-κB pathway to induce primary rat spinal cord astrocytes dedifferentiation

AimsAstrocytes re-acquire stem cell potential upon inflammation, thereby becoming a promising source of cells for regenerative medicine. Nanog is an essential transcription factor to maintain the characteristics of stem cells. We aimed to investigate the role of Nanog in astrocyte dedifferentiation. Main methodsTNF-α was used to induce the dedifferentiation of primary rat spinal cord astrocytes. The expression of immature markers CD44 and Musashi-1 was detected by qRT-PCR and immunofluorescence. The Nanog gene is knocked down by small interference RNA. Nanog expression was measured by qRT-PCR and western blotting. BAY 11–7082 was used to suppress NF-κB signals in astrocytes. NF-κB signaling was evaluated by Western blotting. Key findingsOur results showed that TNF-α promoted the re-expression of CD44 and Musashi-1 in astrocytes. Dedifferentiated astrocytes could be induced to differentiate into oligodendrocyte lineage cells indicating that the astrocytes had pluripotency. In addition, TNF-α treatment activated NF-κB signaling pathway and up-regulated Nanog. Knockdown of Nanog reversed the increase of CD44 and Musashi-1 induced by TNF-α without affecting the activation of NF-κB signaling. Importantly, blocking NF-κB signaling by BAY 11–7082 inhibited the expression of immature markers suggesting that TNF-α induces dedifferentiation of astrocytes through the NF-κB signaling pathway. BAY 11–7082 could also inhibit the expression of Nanog, which indicated that Nanog was regulated by NF-κB signaling pathway. SignificanceThese findings indicate that activation of the NF-κB signaling pathway through TNF-α leads to astrocytes dedifferentiation via Nanog. These results expand our understanding of the mechanism of astrocytes dedifferentiation.

Rejuvenation of Helicobacter pylori–Associated Atrophic Gastritis Through Concerted Actions of Placenta-Derived Mesenchymal Stem Cells Prevented Gastric Cancer

Chronic Helicobacter pylori infection causes gastric cancer via the progression of precancerous chronic atrophic gastritis (CAG). Therefore, repairing gastric atrophy could be a useful strategy in preventing H. pylori–associated gastric carcinogenesis. Although eradication of the bacterial pathogen offers one solution to this association, this study was designed to evaluate an alternative approach using mesenchymal stem cells to treat CAG and prevent carcinogenesis. Here, we used human placenta-derived mesenchymal stem cells (PD-MSCs) and their conditioned medium (CM) to treat H. pylori–associated CAG in a mice/cell model to explore their therapeutic effects and elucidate their molecular mechanisms. We compared the changes in the fecal microbiomes in response to PD-MSC treatments, and chronic H. pylori–infected mice were given ten treatments with PD-MSCs before being sacrificed for end point assays at around 36 weeks of age. These animals presented with significant reductions in the mean body weights of the control group, which were eradicated following PD-MSC treatment (p < 0.01). Significant changes in various pathological parameters including inflammation, gastric atrophy, erosions/ulcers, and dysplastic changes were noted in the control group (p < 0.01), but these were all significantly reduced in the PD-MSC/CM-treated groups. Lgr5+, Ki-67, H+/K+-ATPase, and Musashi-1 expressions were all significantly increased in the treated animals, while inflammatory mediators, MMP, and apoptotic executors were significantly decreased in the PD-MSC group compared to the control group (p < 0.001). Our model showed that H. pylori–initiated, high-salt diet–promoted gastric atrophic gastritis resulted in significant changes in the fecal microbiome at the phylum/genus level and that PD-MSC/CM interventions facilitated a return to more normal microbial communities. In conclusion, administration of PD-MSCs or their conditioned medium may present a novel rejuvenating agent in preventing the progression of H. pylori–associated premalignant lesions.

Musashi-1 Regulates MIF1-Mediated M2 Macrophage Polarization in Promoting Glioblastoma Progression

Simple SummaryGlioblastoma (GBM) is the most lethal type of brain cancer. It is well known that the malignancy of cancers is dependent not only on the oncogenic properties of the tumor cells, but also on the composition of the tumor microenvironment, which includes macrophages of the immune system. The prevalence of M2 type macrophages usually promotes tumor progression as opposed to tumor-suppressing function of M1 type macrophages. In our previous studies, we identified Musashi-1 (MSI1) RNA-binding protein as a principal oncogenic factor in GBM. In this study, in a pursuit of finding secreted factors that may alter tumor microenvironment in GBM, we identified MIF1 cytokine to be positively regulated by MSI1. Moreover, we found that MSI1-mediated MIF1 secretion promotes differentiation of macrophages into pro-oncogenic M2 phenotype. The oncogenic role of MSI1/MIF1/M2 macrophage regulatory axis was also confirmed in GBM mouse models, which makes it a promising target for novel drug discovery.Glioblastoma (GBM) is the most malignant brain tumor which is characterized by high proliferation and migration capacity. The poor survival rate has been attributed to limitations of the current standard therapies. The search for novel biological targets that can effectively hamper tumor progression remains extremely challenging. Previous studies indicated that tumor-associated macrophages (TAMs) are the abundant elements in the tumor microenvironment that are closely implicated in glioma progression and tumor pathogenesis. M2 type TAMs are immunosuppressive and promote GBM proliferation. RNA-binding protein Musashi-1 (MSI1) has recently been identified as a marker of neural stem/progenitor cells, and its high expression has been shown to correlate with the growth of GBM. Nevertheless, the relationship between MSI1 and TAMs in GBM is still unknown. Thus, in our present study, we aimed to investigate the molecular interplay between MSI1 and TAMs in contributing to GBM tumorigenesis. Our data revealed that the secretion of macrophage inhibitory factor 1 (MIF1) is significantly upregulated by MSI1 overexpression in vitro. Importantly, M2 surface markers of THP-1-derived macrophages were induced by recombinant MIF1 and reduced by using MIF1 inhibitor (S,R)-3-(4-hHydroxyphenyl)-4,5-dihydro-5-isoxazole acetic acid (ISO-1). Furthermore, GBM tumor model data suggested that the tumor growth, MIF1 expression and M2 macrophage population were significantly downregulated when MSI1 expression was silenced in vivo. Collectively, our findings identified a novel role of MSI1 in the secretion of MIF1 and the consequent polarization of macrophages into the M2 phenotype in promoting GBM tumor progression.

Expression of Musashi-1 Increases in Bone Healing.

Musashi-1 (MSI1) is an RNA-binding protein that regulates progenitor cells in adult and developing organisms to maintain self-renewal capacities. The role of musashi-1 in the bone healing environment and its relation with other osteogenic factors is unknown. In the current study, we analyze the expression of MSI1 in an experimental model of rat femoral bone fractures. We also analyze the relation between MSI1 expression and the expression of two osteogenic markers: periostin (POSTN) and runt-related transcription factor 2 (RUNX2). We use histological, immunohistochemical, and qPCR techniques to evaluate bone healing and the expression of MSI1, POSTN, and RUNX2 over time (4, 7, and 14 days). We compare our findings with non-fractured controls. We find that in bone calluses, the number of cells expressing MSI1 and RUNX2 increase over time and the intensity of POSTN expression decreases over time. Within bone calluses, we find the presence of MSI1 expression in mesenchymal stromal cells, osteoblasts, and osteocytes but not in hypertrophic chondrocytes. After 14 days, the expression of MSI1, POSTN, and RUNX2 was significantly correlated. Thus, we conclude that musashi-1 potentially serves in the osteogenic differentiation of mesenchymal stromal cells and bone healing. Therefore, further studies are needed to determine the possibility of musashi-1′s role as a clinical biomarker of bone healing and therapeutic agent for bone regeneration.

Regulatory T cells promote glioma cell stemness through TGF-\u03b2\u2013NF-\u03baB\u2013IL6\u2013STAT3 signaling

Glioma stem cells (GSCs) contribute to the malignant growth of glioma, but little is known about the interaction between GSCs and tumor microenvironment. Here, we found that intense infiltration of regulatory T cells (Tregs) facilitated the qualities of GSCs through TGF-β secretion that helped coordinately tumor growth. Mechanistic investigations indicated that TGF-β acted on cancer cells to induce the core cancer stem cell-related genes CD133, SOX2, NESTIN, MUSASHI1 and ALDH1A expression and spheres formation via NF-κB–IL6–STAT3 signaling pathway, resulting in the increased cancer stemness and tumorigenic potential. Furthermore, Tregs promoted glioma tumor growth, and this effect could be abrogated with blockade of IL6 receptor by tocilizumab which also demonstrated certain level of therapeutic efficacy in xenograft model. Additionally, expression levels of CD133, IL6 and TGF-β were found to serve as prognosis markers of glioma patients. Collectively, our findings reveal a new immune-associated mechanism underlying Tregs-induced GSCs. Moreover, efforts to target this network may be an effective strategy for treating glioma.

Increased expression of MUSASHI1 in epithelial breast cancer cells is due to down regulation of miR-125b

BackgroundMusashi1 (MSI1) is an oncogenic protein with a crucial role in the proliferation and characteristics of the epithelial cells in breast cancer. The change in expression of MSI1 has a role in solid tumor progression. There are different factors that regulate MSI1 expression in various cancer tissues including microRNAs which are considered as one of the most important of these factors. The aim of our study is identification of the molecular cause of maximal expression of MSI1 in epithelial breast cancer cell lines.ResultsAmong predicted microRNAs, miR-125b, miR-637 and miR-802 were able to significantly reduce the luciferase activity. In addition, the relative expression of these three miRNAs were measured in the cancerous cell lines that results showed a significant reduction in expression of all microRNAs. On the other hand, only the overexpression of miR-125b caused a change in the expression pattern of MSI1 in breast epithelial cancer cell lines. Accordingly, our results demonstrated that the exogenous expression of miR-125b decreased not only the MSI1 protein but also expression of epithelial markers in breast cancer cells.ConclusionsThe results of luciferase reporter assay showed that MSI1 is a direct target for miR-125b in epithelial breast cancer cells. Moreover, higher amount of MSI1 in those cell lines seems due to the reduced amount of miR-125b, which is responsible for epithelial features of those kinds of cancer cells. Therefore, the modulation of miR-125b may be a potential approach to help to combat against epithelial breast tumors.

Musashi1 expression is negatively correlated with numb expression in brain metastases.

The expression of tumor stem cell markers musashi1 (msi1) and numb in brain metastases were detected to explore their roles in the development of brain metastases.A total of 51 cases of brain metastasis, 29 cases of primary tumor and 15 cases of normal brain tissue were selected. Immunohistochemistry and reverse transcription polymerase chain reaction (RT-PCR) were used to detect msi1 and numb expression at the protein and mRNA levels. Correlation between msi1 and numb in brain metastases were evaluated.Immunohistochemistry and RT-PCR showed that no significant difference in the expression of msi1 and numb between brain metastases and primary tumors was observed (P > .05); the expression of msi1 and numb in brain metastases was significantly higher than that in normal brain tissues (P < .05); and the expression of msi1 and numb in primary tumors was significantly higher than that in normal brain tissues (P < .05). In general, the expression of msi1 gene was negatively correlated with the expression of numb at mRNA level by Pearson correlation analysis (r = −0.345, P < .05). Additionally, the expression of msi1 and numb in brain metastases was not related to gender, age, and tissue origin (P > .05).Msi1 is highly expressed in brain metastases and primary tumors, while numb is lowly expressed in brain metastases and primary tumors; msi1 and numb are negatively correlated in brain metastases, suggesting that msi1 and numb may have regulatory mechanisms in the development of brain metastases.

Long noncoding RNA LEF1‐AS1 acts as a microRNA‐10a‐5p regulator to enhance MSI1 expression and promote chemoresistance in hepatocellular carcinoma cells through activating AKT signaling pathway

Long noncoding RNAs (lncRNAs) contribute to the development of hepatocellular carcinoma (HCC), which could regulate various HCC biological characteristics. Here, the study seeks to investigate the role of lncRNA LEF1-AS1 in HCC cell chemoresistance by regulating microRNA (miR)-10a-5p and Musashi1 (MSI1). The microarray-based analysis was employed to identify the HCC-related lncRNA-miRNA-gene regulatory network. Expression patterns of LEF1-AS1, miR-10a-5p, and MSI1 in the HCC cell lines, tissues were accessed by means of reverse transcription-quantitative polymerase chain reaction. Next, the interaction among LEF1-AS1, miR-10a-5p, and MSI1 in HCC was accessed by bioinformatics and dual-luciferase reporter gene assay. Then, the cell line resistant to cisplatin was established, which was then treated with sh/oe-lncRNA LEF1-AS1, miR-10a-5p-mimic, and oe/sh-MSI1 vectors alone or in combination. Afterward, the effect of LEF1-AS1, miR-10a-5p, and MSI1 on HCC cell chemoresistance, proliferation, and apoptosis was assessed. At last, in vivo experiments confirmed the role of MSI1 in tumor growth and chemoresistance in HCC. LEF1-AS1 might potentially affect the growth and chemoresistance of HCC cells by regulating miR-10a-5p and MSI1. LEF1-AS1 and MSI1 expression patterns were elevated, while miR-10a-5p was repressed in HCC tissues and cell lines. LEF1-AS1 combined to miR-10a-5p and regulated MSI1, thereby activating the protein kinase B (AKT) signaling pathway. Knockdown of LEF1-AS1 and MSI1 or elevation of miR-10a-5p compromised the proliferation of Huh7 cell line resistant to DDP and promoted its chemosensitivity and apoptosis. At last, these in vitro findings were also confirmed in vivo. Our results unraveled LEF1-AS1 acts as a miR-10a-5p modulator to promote chemoresistance of HCC cells by stimulating MSI1 and activating the AKT signaling pathway, which might provide a novel therapeutic target for HCC.

Interaction between SDF1 and CXCR4 Promotes Photoreceptor Differentiation via Upregulation of NFκB Pathway Signaling Activity in Pax6 Gene-Transfected Photoreceptor Precursors

Background: CCL2 (also known as monocyte chemoattractant protein 1) and CX3CR1 (also known as Fractalkine receptor)-deficient mice have damaged photoreceptors. Objectives: We examined the interaction of SDF1 and CXCR4 on the differentiation of retinal progenitors into rhodopsin-positive photoreceptors. Methods: Cloned retinal progenitors were obtained by Pax6 gene transfection of mouse iPS cells followed by serial dilution. Clones were selected by expression of nestin, Musashi1, Six3, and Chx10 mRNA. Cell surface protein expression was analyzed by flow cytometry. The levels of mRNA and intracellular protein were examined by real-time PCR and immunochemistry, respectively. Transient transfection experiments of retinal progenitors were conducted using a human rhodopsin promoter luciferase plasmid. Results: We selected 10 clones that expressed Six3, Chx10, Crx, Rx1, Nrl, CD73, and rhodopsin mRNA, which, except for rhodopsin, are photoreceptor precursor markers. Clones expressed both CD73 and CXCR4 on the cell surface and differentiated into rhodopsin-positive photoreceptors, which was reinforced by the addition of exogenous SDF1. A CXCR4 inhibitor AMD3100 blocked SDF1-mediated differentiation of progenitors into photoreceptors. SDF1 enhanced human rhodopsin promoter transcription activity, possibly via the NFκB pathway. Addition of SDF1 to the cell culture induced nuclear translocation of NFκB on retinal progenitor cell clones. Neonatal and newborn mouse retinas expressed SDF1 and CXCR4. Cells in the outer nuclear layer where photoreceptors are located expressed CXCR4 at P14 and P56. Cells in the inner nuclear layer expressed SDF1. Conclusions: These findings suggest that retinal progenitor cell differentiation was at least partly regulated by SDF1 and CXCR4 via upregulation of NFκB activity.